Copper(II) is known to bind in the influenza virus His37 cluster in the homotetrameric M2 proton channel and block the proton current needed for uncoating. Copper complexes based on iminodiacetate also block the M2 proton channel and show reduced cytotoxicity and zebrafish-embryo toxicity. In voltage-clamp oocyte studies using the ubiquitous amantadine-insensitive M2 S31N variant, the current block showed fast and slow phases in contrast to the single phase found for amantadine block of WT M2. Here we evaluate the mechanism of block by copper adamantyl iminodiacitate (Cu(AMT-IDA)) and copper cyclooctyl iminodiacitate (Cu(CO-IDA)) complexes and address whether the complexes can coordinate with one or more of the His37 imidazoles. The current traces were fitted to parametrized master equations. The energetics of binding and the rate constants suggest that the first step is copper-complex binding within the channel and the slow step in the current block is the formation of a Cu-histidine coordination complex. Solution-phase isothermal titration calorimetry (ITC) and density functional theory (DFT) calculations indicate that imidazole binds to the copper complexes. Structural optimization using DFT reveals that the complexes fit inside the channel and project the Cu(II) towards the His37 cluster allowing one imidazole to form a covalent bond with the Cu(II). Electrophysiology and DFT studies also show that the complexes block the G34E amantadine-resistant mutant in spite of some crowding in the binding site by the glutamates.

中文翻译：

---

铜配合物与甲型流感 M2 S31N 和 S31N/G34E 通道结合的机制和动力学

已知铜 (II) 在同四聚体 M2 质子通道中的流感病毒 His37 簇中结合并阻断脱壳所需的质子电流。基于亚氨基二乙酸盐的铜配合物也阻断 M2 质子通道，并显示出降低的细胞毒性和斑马鱼胚胎毒性。在使用普遍存在的金刚烷胺不敏感 M2 S31N 变体的电压钳卵母细胞研究中，与 WT M2 的金刚烷胺阻滞发现的单相相比，电流阻滞显示出快相和慢相。在这里，我们评估了金刚烷基亚氨基二酸铜 (Cu(AMT-IDA)) 和环辛基亚氨基二酸铜 (Cu(CO-IDA)) 配合物的阻断机制，并讨论了这些配合物是否可以与一种或多种 His37 咪唑配位。电流轨迹适合参数化的主方程。结合的能量学和速率常数表明，第一步是通道内的铜复合物结合，当前块中的缓慢步骤是铜-组氨酸配位复合物的形成。溶液相等温滴定量热法 (ITC) 和密度泛函理论 (DFT) 计算表明咪唑与铜配合物结合。使用 DFT 进行的结构优化表明，配合物适合通道内并将 Cu(II) 投射到 His37 簇，从而允许一个咪唑与 Cu(II) 形成共价键。电生理学和 DFT 研究还表明，尽管谷氨酸盐在结合位点有一些拥挤，但复合物阻断了 G34E 金刚烷胺抗性突变体。